Gelatin is a protein derived from the connective tissues of vertebrates, that is, collagen. It is produced when collagen is boiled in water. Gelatin hydrolysis detects the presence of gelatinases. Gelatinases are proteases secreted extracellularly by some bacteria which hydrolyze or digest gelatin. The production of gelatinases is used as a presumptive test for the identification of various organisms, including Staphylococcus sp., Enterobacteriaceae, and some gram-positive bacilli.

Principle

This test is used to determine the ability of an organism to produce extracellular proteolytic enzymes (gelatinases) that liquefy gelatin, a component of vertebrate connective tissue.

The reaction occurs in two sequential steps: in first reaction gelatinases hydrolyze gelatin into polypeptides and then polypeptides are further converted into amino acids. The amino acid is taken up by the cell and used for metabolic purposes.

The presence of gelatinases is detected using a nutrient gelatin medium. When an organism produces gelatinase, the enzyme liquefies the growth medium by hydrolyzing gelatin present in the medium.

In some cases, plates are flooded with saturated ammonium sulfate to precipitate unhydrolyzed gelatin, making the clear zones easier to see. Results are often observed within 5 to 10 minutes after flooding with saturated ammonium sulfate.

Expected Results

Positive: Partial or total liquefaction of the inoculated tube (the control tube must be completely solidified) at 4°C within 14 days. On plates, gelatin hydrolysis is indicated by clear zones around gelatinase-positive colonies.

Negative: Complete solidification of the tube at 4°C. On plates, no clear zones around colonies are observed.